These studies examine the effects of methamphetamine (METH) in an in vitro model of rat fetal mesencephalic cells. In addition, we sought to determine if production of nitric oxide was involved in the neurotoxic events associated with the presence of METH in the culture medium. In mesencephalic primary DA cultures, METH (1.5 mM) caused a significant reduction of neurons. Tyrosine hydroxylase (TH) immunohistochemistry showed a marked reduction of TH-positive cells with normal architecture. METH-induced toxic effects were significantly attenuated in the presence of inhibitors of nitric oxide synthase (NOS). Moreover, inhibitors of ADP-ribosylation such as benzamide and nicotinamide also blocks METH toxic effects in vitro. There was an associated increase in reactive gliosis in the presence of methamphetamine. reactive gliosis was also reduced in the presence of these inhibitors. These results suggest that NO formation might play an important role in the manifestation of METH-induced neurotoxicity. When taken together with our in vivo results using the superoxide dismutase transgenic mice, our data further implicate a role for oxidative stress in the manifestation of the deleterious effects of the amphetamines.